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Glycosaminoglycan–Protein Interaction Studies Using Fluorescence Spectroscopy

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Glycosaminoglycans

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1229))

Abstract

Fluorescence spectroscopy is a quantitative analytical tool that has been extensively used to provide structural and dynamical information on GAG–protein complexes. It possesses major advantages including high sensitivity, relative ease of applicability, and wide range of available fluorescence labels and probes. It has been applied to practically every protein–GAG system through the use of either intrinsic (e.g., Trp) or extrinsic (e.g., a non-covalent fluorophore) probe. For studies involving GAGs, it forms the basis for measurement of dissociation constant of complexes and the stoichiometry of binding, which helps elucidate many other thermodynamic and/or mechanistic parameters. We describe the step-by-step procedure to measure the affinity of GAG–protein complexes, parse the ionic and nonionic components of the free energy of binding, and identify the site of GAG binding through competitive binding experiments.

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Acknowledgements

This work was supported by the grants HL090586 and HL107152 from the National Institutes of Health.

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Authors and Affiliations

  1. Department of Medicinal Chemistry and Institute for Structural Biology and Drug Discovery, Virginia Commonwealth University, 800 E. Leigh Street, Suite 212, Richmond, VA, 23219, USA

    Rio S. Boothello, Rami A. Al-Horani & Umesh R. Desai

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  1. Rio S. Boothello
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  2. Rami A. Al-Horani
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  3. Umesh R. Desai
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Corresponding author

Correspondence to Umesh R. Desai .

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Editors and Affiliations

  1. Department of Medicinal Chemistry, University of Utah, Salt Lake City, Utah, USA

    Kuberan Balagurunathan

  2. Department of Genetics, Cell Biology and Development, The University of Minnesota, Minneapolis, Minnesota, USA

    Hiroshi Nakato

  3. Department of Medicinal Chemistry, Virginia Commonwealth University, Richmond, Virginia, USA

    Umesh R. Desai

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Boothello, R.S., Al-Horani, R.A., Desai, U.R. (2015). Glycosaminoglycan–Protein Interaction Studies Using Fluorescence Spectroscopy. In: Balagurunathan, K., Nakato, H., Desai, U. (eds) Glycosaminoglycans. Methods in Molecular Biology, vol 1229. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1714-3_27

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  • DOI: https://doi.org/10.1007/978-1-4939-1714-3_27

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1713-6

  • Online ISBN: 978-1-4939-1714-3

  • eBook Packages: Springer Protocols

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